摘要
目的研究轮状病毒（RV）感染对表达钠/氢交换因子3（NHE3）的Caco-2细胞表面NHE3蛋白水平和生物活性的影响及其调控机制。方法构建表达NHE3的Caco-2细胞模型，分为对照组（CTL组）、RV组、Cdc42拮抗剂Pirl-1组和Pirl-1＋RV组。BCECF-AM法和细胞表面NHE3生物素化法测定Na＋-H＋交换活性和细胞表面NHE3蛋白水平，Western blot法测定细胞Cdc42蛋白水平，免疫共沉淀法检测Cdc42与NHE3的相互作用。结果与CTL组相比，RV感染可引起细胞Na＋-H＋交换活性以及细胞表面NHE3蛋白水平明显降低，此抑制作用可被Cdc42拮抗剂Pirl-1拮抗，此外RV感染可引起Cdc42蛋白水平升高，并且也可增加Cdc42蛋白与NHE3蛋白的结合，这些作用也可被Pirl-1拮抗。结论RV感染可能通过Cdc42依赖性内吞途径调控NHE3蛋白水平及生物活性。
ObjectiveTo observe the effects and regulatory mechanism of rotavirus infection on the expression and bioactivity of Na＋ /H＋ exchanger 3 （NHE3） on Caco-2 cells.MethodsA cell model of Caco-2 cells expressing NHE3 was constructed. Four groups were set up, which were control （CTL） group, rotavirus（RV） infection group, Cdc42 inhibitor （Pirl-1） group and Pirl-1＋ RV group. Bioactivity and expression of NHE3 on the surface of Caco-2 cells were determined by BCECF-AM and biotinylation method, respectively. Expression of Cdc42 protein was measured by Western blot. Co-immunoprecipitation was performed to detect the interaction between NHE3 and Cdc42.ResultsCompared with the CTL group, RV infection significantly inhibited the bioactivity and expression of NHE3 on Caco-2 cells. These inhibitory effects were antagonized by Pirl-1. Moreover, RV infection enhanced the expression of Cdc42 protein and promoted the interaction between NHE3 and Cdc42, which were also antagonized by Pirl-1.ConclusionRV infection might regulate the expression and bioactivity of NHE3 through Cdc42-dependent endocytosis pathway.
作者
牛美兰
王鹏
李玲
陈长英
冯荣芳
陈梓潇
焦佳伟
李媛媛
许昊宇
Niu Meilan, Wang Peng, Li Ling, Chen Changying, Feng Rongfang, Chen Zixiao, Jiao Jiawei, Li Yuanyuan, Xu Haoyu ）(Pharmacology Department, Huanghe Science and Technology College, Zhengzhou 450063, China ; Basic Department, the Nursing College of Zhengzhou University, Zhengzhou 450052, China ; Department of Palliative Care and Hospice Care, the Ninth People＇s Hospital of Zhengzhou, Zhengzhou, 450052, Chin)
出处
《中华微生物学和免疫学杂志》
CAS
CSCD
北大核心
2018年第3期181-186,共6页
Chinese Journal of Microbiology and Immunology
基金
国家自然科学基金（81500433）
河南省教育厅（16A310003,16A320009）
河南省科技厅项目（122300410338,132102310138,142102310140）
郑州市科技攻关项目（2PPTSF302）
